The main theme of my research laboratory is to examine sex-specific epigenetic and circuit changes underlying opioid use disorder. There are three primary questions that we intend to examine under this theme.
1) Identify the role of dorsal striatal projections underlying opioid-induced behavioral plasticity. The striatum forms a critical neural substrate underlying habitual behaviors and is often implicated in reinforcement learning and the development of compulsive habits, a behavioral characteristic of opioid addiction. Habit formation has mostly been attributed to the dorsal striatum and time-dependent intensification of habit formation is thought to occur due to neuroadaptations in the dorsal striatum. Dorsal striatal sub-structures (dorsolateral striatum-DLS and dorsomedial striatum-DMS) receive inputs and send projections from several essential brain areas. Selecting and generating appropriate actions for the execution of behavior involves integrating associative and sensory information. The dorsal striatum and its connections form a key hub where such integration occurs. We will examine how projections to and out of DLS and DMS mediate opioid-induced behavioral plasticity. 2) How does controllable versus uncontrollable stress influence opioid relapse? Stress plays an important role in drug use and relapse. Specifically, daily life stress during abstinence is linked to an increased craving for heroin in individuals with opioid use disorder, which is predictive of a higher risk of relapse. Most importantly, emotional response to a stressful experience is an important determinant of behavioral adaptation to future stressful events. Modulation of stress responses to stressful experiences is a common pathology observed in several psychiatric diseases. The control dimension has most often been examined by using an escapable (controllable) versus non-escapable (uncontrollable) foot shock paradigm. While it is well established that the dorsal striatum mediates stimulus-response learning, the role of the dorsal striatum in influencing freezing or escape responses to aversive stimuli (foot shock) is less understood. We will examine how projections in dorsal striatal substructures (DMS and DLS) modulate aversive learning in a controllable versus non-controllable stress behavioral paradigm, and how pairing such stressors influence drug-taking and relapse assays. Sex differences are an essential determinant of neuroadaptations underlying addiction and stress mechanisms and warrant further exploration. Ovarian hormones regulate dorsal striatal plasticity through the receptors present in the neurons. We will therefore examine questions 1 and 2 between males and females. We will utilize retrograde tracing followed by immunofluorescence and in-situ-hybridization to characterize projections to and out of DS substructures in opioid addiction and or stress: We will then them genetically activate or inhibit these circuits to examine their role in these behaviors. Finally, we will deplete ovarian hormones in females through ovariectomy and examine the sex-specific circuit plasticity underlying stress and or heroin exposure. 3) Identify the neuronal circuit-specific transcriptional and epigenetic changes that contribute to maladaptive plasticity during opioid relapse and or stress. Opioids are thought to alter transcriptional and chromatin processes in brain reward regions modulating synaptic plasticity underlying the addicted state. Experiences and cues associated with learning have been observed to induce distinct neural gene expression patterns in a temporal and pulsative manner, which is analogously referred to as the genomic action potential (gAP). Mechanistically, gAP has cascading consequences that influence long-term functioning required for behavioral adaptations, which is achieved through governing of sex and cell-type-specific transcriptome and the translatome. Such gAP-mediated transcriptional and translational changes over a period of time enable experience-dependent modification of the synaptic circuitry to catalyze behavioral adaptations. There is emerging evidence that gAP underlies experience-dependent neuroplasticity through immediate early genes (IEG). However, the characteristics and functions of such neuroadaptations within specific neural substrates and between sexes in stress and opioid-induced plasticity remain unclear. We will examine time-dependent changes in IEGs in the DS in rodents that will undergo heroin self-administration, relapse and stress assays. This will be addressed through biochemical approaches, including RNAscope, spatial transcriptomics, and CatFish. To determine cell-type specific (i.e. the D1 and D2 expressing medium spiny neurons) IEG changes in the DMS and DLS, we will utilize RiboTag capture using Cre-inducible RiboTag in transgenic D1 and D2 cre lines. RiboTag will allow the isolation of ribosomal RNA for sex and cell-type-specific quantification of IEGs. We will then do viral manipulation of IEGs in specific cell types in DMS and DLS. This will enable dissecting the role of D1 and D2 MSN in the DMS and DLS in stress, drug-taking, and relapse. To examine the patterns of genomic enrichment of IEGs, we will use Chromatin immunoprecipitation (ChIP) followed by RNA-seq to examine patterns of gene expression that may be related to IEG binding.
Swarup Mitra, PhD